ethyl N2-dodecanoyl-.sc.l.sc.-argininate hydrochloride

Acute Toxicity: inhalation

Administrative data

Endpoint:

acute toxicity: inhalation

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From November 17, 2005 to March 17, 2006

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Justification for type of information:

Guideline study with GLP.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2006

Report date:

2006

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 403 (Acute Inhalation Toxicity)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.2 (Acute Toxicity (Inhalation))

Deviations:

no

Test guideline 3

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.1300 (Acute inhalation toxicity)

Deviations:

no

GLP compliance:

yes

Test type:

standard acute method

Limit test:

yes

Test material

Test material information

Constituent 1

Reference substance name:

-

EC Number:

434-630-6

EC Name:

-

Cas Number:

60372-77-2

Molecular formula:

Hill formula: C20H41N4O3Cl

IUPAC Name:

ethyl N2-dodecanoyl-l-argininate hydrochloride

Details on test material:

- Name of test material: LAE in ethanol
- Physical state: Spray can
- Lot/batch No.: LI-531
- Storage condition of test material: Room temperature / darkness

Specific details on test material used for the study:

Substance: L.A.E. in ethanol
Batch number: LI-631 as of Oct. 19, 2005, content 0.634% of LAE

Test animals

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Adult SPF bred Wistar Unilever rats, strain Hsd Cpb:WU from the experimental breeder Harlan-Winkelmann GmbH, Borchen (Germany) and Harlan Nederland (The Netherlands)
- Age at study initiation: 2 months old
- Weight at study initiation: Variation of individual weight did not exceed ±10% of the mean for each sex. Females: 173-187 g; males 176-201 g
- Housing: Singly in conventional Makrolon® Type IIIH cages
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: 5 days prior to start

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22±2ºC
- Humidity (%): 40-70%
- Air changes (per hr): 10 air changes per hour approximately
- Photoperiod (hrs dark / hrs light): 12 hours artificial light each 24 hours

IN-LIFE DATES: No mortalities

Administration / exposure

Route of administration:

inhalation: aerosol

Type of inhalation exposure:

nose only

Vehicle:

air

Details on inhalation exposure:

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: Exposure tubes applying a directed-flow nose-only exposure principle.
- Exposure chamber volume: 50 L
- Source and rate of air: 20 l/min
- Method of conditioning air: Calibrated flowmeter/Soap bubble meter
- System of generating particulates/aerosols: Magnetic piston equipped with two electronic clocks
- Method of particle size determination: TSI-Laser velocimeter APS
- Treatment of exhaust air: Exhaust air was purified via cotton-wool/HEPA filters
- Temperature, humidity, pressure in air chamber: 21.9-23.2 ºC, humidity values were lower than recommended due to the use of conditioned, dry air for dispersion of the test substance.

TEST ATMOSPHERE
- Brief description of analytical method used: Gas chromatographic method (for propellants); HPLC (for the active ingredient)
- Samples taken from breathing zone: yes, in the vicinity of the breathing zone of the animals

VEHICLE
- Composition of vehicle (if applicable): Undiluted

TEST ATMOSPHERE (if not tabulated)
- Particle size distribution: Adequate to reach all potential target structures of the respiratory tract
- MMAD (Mass median aerodynamic diameter) / GSD (Geometric st. dev.): 4.6 um / 1.92

Analytical verification of test atmosphere concentrations:

yes

Duration of exposure:

4 h

Concentrations:

Target concentration: 25000 mg/m3 (nominal 42000mg/m3).

No. of animals per sex per dose:

Group 1: Target concentration 0 mg/m3
5 male per dose
5 female per dose

Group 2: Target concentration: 25.000 mg/m3
5 male per dose
5 female per dose

Control animals:

yes

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: Weighing record: before exposure, day 3, 7 and 14; Observations: Several times on the day of exposure, at least once daily thereafter.
- Necropsy of survivors performed: yes
- Other examinations performed: clinical signs, body weight and reflexes.

Statistics:

Statistical Evaluation of data:

Necropsy findings: In case of findings: Fisher test
Body weights: One-way ANOVA (vida infra)
Physiological data: Data of rectal temperature measurements are statistically evaluated using the ANOVA procedure (vida infra)
LC50: Rosello et al.; Schaper et al.
Analysis of variance: ANOVA (p= 0.05)

Results and discussion

Effect levels

Effect levels 1

Key result

Sex:

male/female

Dose descriptor:

LC50

Effect level:

> 28 150 mg/m³ air

Based on:

test mat.

Exp. duration:

4 h

Remarks on result:

other: LAE in ethanol

Effect levels 2

Key result

Sex:

male/female

Dose descriptor:

LC50

Effect level:

> 5 883 mg/m³ air (analytical)

Based on:

act. ingr.

Exp. duration:

4 h

Remarks on result:

other: LAE

Mortality:

Mortality was not observed.

Clinical signs:

other: Group 1: All rats tolerated the exposure without specific signs. Group 2/Males: Bradypnea, labored breathing patterns, irregular breathing patterns, high-legged gait, piloerection. Group 2/Females: Labored breathing patterns, irregular breathing patterns,

Body weight:

Comparisons did not reveal changes in body weights considered to be of toxicological significance.

Gross pathology:

Animals sacrified at the end of the observation period: The macroscopic findings were indistinguishable amongst the groups.

Any other information on results incl. tables

Results obtained after exposure of rats for 4 hours:

Group/sex	Target concentration (mg/m^3)	Toxicological result(*)	Onset and duration signs	Onset mortality
1 / m	0	0/0/5	-	-
2 / m	25000	0/4/5	0 d - 3 d	-
1 / f	0	0/0/5	-	-
2 / f	25000	0/1/5	0 d - 4 d	-

(*) Number of dead animals/Number of animals with signs after cessation of exposure/Number of animals exposed.

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Remarks:

CLP Implementation.

Conclusions:

Combined experimental evidence suggest that the test substance has a mild respiratory tract irritation potential if exposure is sufficiently high. Thus, the LC50 is greater than 28150 mg/m3 (LAE in ethanol). The LC50 for LAE is greater than 5883 mg/m3 (analytical concentration).

Executive summary:

The test conditions applied fulfilled recommendations of current test guidelines according to the OECD Guideline Nº 403 and EU method B.2, with regard to attainment of steady-state concentration, respirability of particles and the limit concentration tested. The aerosolized test substance proved to have virtually no acute inhalation toxicity to rats and mortality did not occurred even at levels of exposure exceeding significantly the limit concentration called for by the current testing guidelines applied. Combined experimental evidence, however, would suggest that the test substance has a mild respiratory tract irritation potential, if exposure to the aerosol is sufficiently high. Thus, for the test substance the LC₅₀ is greater than 28150 mg/m³ (LAE in ethanol). The LC50 for LAE is greater than 5883 mg/m³ (analytical concentration).